Role Of GRHL2 Research Articles

Mild overexpression of a novel transcription factor Grainy head like-2 (Grhl2) elicits hybrid phenotype in human astrocytoma cells

The Grainy head like-2 (Grhl2) transcription factor plays a major role in embryonic and cancer development. The role of Grhl2 has been intensively studied in various cancers but not for brain cancer. Hence, in this study, we provide a preliminary understanding on the role of Grhl2 that regulate the transition of astrocytoma cells. The human A172 astrocytoma cell line, a mesenchymal cell characterized by mild overexpression of Grhl2 transcription factor, was used in this study. At first, the Grhl2 stably overexpressing A172 clones into three types i.e., Grhl2+ (mild), Grhl2++ (moderate) and Grhl2+++ (high) based on mRNA and protein expression levels of Grhl2 were characterized. Phenotypic characteristics of vector and Grhl2+ cells were observed using phase contrast microscopy. Quantitative PCR (qPCR), Western blot and immunofluorescence were used to detect the level of mesenchymal markers (N-cadherin/vimentin) and also epithelial markers (E-cadherin/ β- catenin) in vector and Grhl2+ cells. The migration and invasion characteristics of vector and Grhl2+ cells were determined by scratch assay and Boyden chamber assay. Further, the Grhl2+ cells were characterized to determine the effect of temozolomide chemotherapy drug which were widely used in treating brain cancer. As expected, in phase contrast image, we observed the mesenchymal characteristic of A172 cells becomes hybrid phenotype i.e., mixture of mesenchymal (spindle-like fibroblast morphology) and epithelial (cobblestone like appearance) cells upon Grhl2 mild expression (Grhl2+) when compared to vector cells. Further, we found that there was a significant upregulation of E-cadherin at both mRNA and protein levels in Grhl2+ cells when compared to vector cells. There was a significant upregulation of β-catenin, N-cadherin and vimentin at mRNA levels, but there was no significant upregulation at the protein levels in Grhl2+ cells compared to the vector cells. The migration and invasion were diminished in Grhl2+ cells when compared to the vector control cells. We observed that the Grhl2+ were sensitive to the temozolomide compared to the vector cells. This infers that the Grhl2+ cells were unable to attain complete transition of mesenchymal to epithelial state, and hence we categorized the Grhl2+ cells as hybrid phenotype. The results provide a better understanding of the largely unknown function of Grhl2 in human astrocytoma cells as tumor progression or suppression.

VULCAN integrates ChIP-seq with patient-derived co-expression networks to identify GRHL2 as a key co-regulator of ERa at enhancers in breast cancer

BackgroundVirtUaL ChIP-seq Analysis through Networks (VULCAN) infers regulatory interactions of transcription factors by overlaying networks generated from publicly available tumor expression data onto ChIP-seq data. We apply our method to dissect the regulation of estrogen receptor-alpha activation in breast cancer to identify potential co-regulators of the estrogen receptor’s transcriptional response.ResultsVULCAN analysis of estrogen receptor activation in breast cancer highlights the key components of the estrogen receptor complex alongside a novel interaction with GRHL2. We demonstrate that GRHL2 is recruited to a subset of estrogen receptor binding sites and regulates transcriptional output, as evidenced by changes in estrogen receptor-associated eRNA expression and stronger estrogen receptor binding at active enhancers after GRHL2 knockdown.ConclusionsOur findings provide new insight into the role of GRHL2 in regulating eRNA transcription as part of estrogen receptor signaling. These results demonstrate VULCAN, available from Bioconductor, as a powerful predictive tool.

Grainyhead-like Protein 2: The Emerging Role in Hormone-Dependent Cancers and Epigenetics.

In mammals, the grainyhead-like transcription factor (GRHL) family is composed of three nuclear proteins that are responsible for driving epithelial cell fate: GRHL1, GRHL2, and GRHL3. GRHL2 is important in maintaining proper tubulogenesis during development and in suppressing the epithelial-to-mesenchymal transition. Within the last decade, evidence indicates both tumor-suppressive and oncogenic roles for GRHL2 in various types of cancers. Recent studies suggest that GRHL2 may be especially important in hormone-dependent cancers, as correlative relationships exist between GRHL2 and various steroid receptors, such as the androgen and estrogen receptors. Acting as a pioneer factor and coactivator, GRHL2 may directly affect steroid receptor transcriptional activity. This review will highlight recent discoveries of GRHL2 activity in cancer and in maintaining the epithelial state, while also exploring recent literature on the role of GRHL2 in hormone-dependent cancers and epigenetics.

Mutations in GRHL2 Result in an Autosomal-Recessive Ectodermal Dysplasia Syndrome

Grainyhead-like 2, encoded by GRHL2, is a member of a highly conserved family of transcription factors that play essential roles during epithelial development. Haploinsufficiency for GRHL2 has been implicated in autosomal-dominant deafness, but mutations have not yet been associated with any skin pathology. We investigated two unrelated Kuwaiti families in which a total of six individuals have had lifelong ectodermal defects. The clinical features comprised nail dystrophy or nail loss, marginal palmoplantar keratoderma, hypodontia, enamel hypoplasia, oral hyperpigmentation, and dysphagia. In addition, three individuals had sensorineural deafness, and three had bronchial asthma. Taken together, the features were consistent with an unusual autosomal-recessive ectodermal dysplasia syndrome. Because of consanguinity in both families, we used whole-exome sequencing to search for novel homozygous DNA variants and found GRHL2 mutations common to both families: affected subjects in one family were homozygous for c.1192T>C (p.Tyr398His) in exon 9, and subjects in the other family were homozygous for c.1445T>A (p.Ile482Lys) in exon 11. Immortalized keratinocytes (p.Ile482Lys) showed altered cell morphology, impaired tight junctions, adhesion defects, and cytoplasmic translocation of GRHL2. Whole-skin transcriptomic analysis (p.Ile482Lys) disclosed changes in genes implicated in networks of cell-cell and cell-matrix adhesion. Our clinical findings of an autosomal-recessive ectodermal dysplasia syndrome provide insight into the role of GRHL2 in skin development, homeostasis, and human disease.

Evidence for multiple roles for grainyhead-like 2 in the establishment and maintenance of human mucociliary airway epithelium

Most of the airways of the human lung are lined by an epithelium made up of ciliated and secretory luminal cells and undifferentiated basal progenitor cells. The integrity of this epithelium and its ability to act as a selective barrier are critical for normal lung function. In other epithelia, there is evidence that transcription factors of the evolutionarily conserved grainyheadlike (GRHL) family play key roles in coordinating multiple cellular processes required for epithelial morphogenesis, differentiation, remodeling, and repair. However, only a few target genes have been identified, and little is known about GRHL function in the adult lung. Here we focus on the role of GRHL2 in primary human bronchial epithelial cells, both as undifferentiated progenitors and as they differentiate in air-liquid interface culture into an organized mucociliary epithelium with transepithelial resistance. Using a dominant-negative protein or shRNA to inhibit GRHL2, we follow changes in epithelial phenotype and gene transcription using RNA sequencing or microarray analysis. We identify several hundreds of genes that are directly or indirectly regulated by GRHL2 in both undifferentiated cells and air-liquid interface cultures. Using ChIP sequencing to map sites of GRHL2 binding in the basal cells, we identify 7,687 potential primary targets and confirm that GRHL2 binding is strongly enriched near GRHL2-regulated genes. Taken together, the results support the hypothesis that GRHL2 plays a key role in regulating many physiological functions of human airway epithelium, including those involving cell morphogenesis, adhesion, and motility.

Correction: Grhl2 Determines the Epithelial Phenotype of Breast Cancers and Promotes Tumor Progression

[This corrects the article on p. e50781 in vol. 7.].